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The present investigation was undertaken to assess genetic diversity in seventy genotypes of kodo millet. The analysis of variance revealed the presence of significant variation among the genotypes for all 13 characters. Higher genotypic coefficient of variation (GCV), phenotypic coefficient of variation (PCV), heritability and genetic advance as per cent mean were recorded for thumb raceme length, number of productive tillers per plant, length of panicle, raceme length and grain yield per plant, indicating that simple selection could be practised for improving these traits. Seventy genotypes were grouped into seven different clusters on the basis of magnitude of D2 values by Mahalanobis D2 analysis. Cluster I had 51 genotypes followed by cluster II with 14 genotypes, while clusters III, IV, V, VI and VII were mono genotypic. The inter–cluster distance was high between clusters II and III and therefore it is suggested to use these genotypes as parents for hybridization to evolve potential segregants.

Mycobacterium abscessus pulmonary infections are increasingly problematic, especially for immunocompromised individuals and those with underlying lung conditions. Currently, there is no reliable standardized treatment, underscoring the need for improved preclinical drug testing. We present a simplified immunosuppressed mouse model using only four injections of cyclophosphamide, which allows for sustained M. abscessus lung burden for up to 16 days. This model proved effective for antibiotic efficacy evaluation, as demonstrated with imipenem or amikacin., Competing Interests: The authors declare no conflict of interest.

CD4+ regulatory T cells (Tregs) are key orchestrators of the immune system, fostering the establishment of protective immunity while preventing deleterious responses. Infancy and childhood are crucial periods of rapid immunologic development, but how Tregs mediate immune responses at these earliest timepoints of human life is poorly understood. In this study, we compare blood and tissue (tonsil) Tregs across pediatric and adult subjects to investigate age-related differences in Treg biology. We observed increased FOXP3 expression and proportions of Tregs in tonsil compared with paired blood samples in children. Within tonsil, early life Tregs accumulated in extrafollicular regions with cellular interactions biased toward CD8+ T cells. Tonsil Tregs in both children and adults expressed transcriptional profiles enriched for lineage defining signatures and canonical functionality compared with blood, suggesting tissue as the primary site of Treg activity. Early life tonsil Tregs transcriptional profiles were further defined by pathways associated with activation, proliferation, and polyfunctionality. Observed differences in pediatric tonsil Treg transcriptional signatures were associated with phenotypic differences, high proliferative capacity, and robust production of IL-10 compared with adult Tregs. These results identify tissue as a major driver of Treg identity, provide new insights into developmental differences in Treg biology across the human lifespan, and demonstrate unique functional properties of early life Tregs., (Copyright © 2024 by The American Association of Immunologists, Inc.)

The establishment of memory T cell responses is critical to protection against pathogens and is influenced by the conditions under which memory formation occurs. Iron is an essential micronutrient for multiple immunologic processes and nutritional deficiency is a common problem worldwide. Despite its prevalence, the impact of nutritional iron deficiency on the establishment of memory T cell responses is not fully understood. In this study we investigate the impact of nutritional iron deficiency on the generation, phenotype, and function of memory T cell responses using a murine model of dietary iron modulation in the context of influenza infection. Iron deficient mice have decreased systemic iron levels and develop significant anemia. Increased T cell expression of the transferrin receptor (CD71) is seen in iron deficient mice at baseline. During primary influenza infection, iron deficient mice experience increased weight loss and phenotypic evidence of impairments in T cell activation. Following recovery from infection, iron deficient mice generate increased influenza specific memory T cells which exhibit impaired ability to produce IFNγ, most notably within the lung. Importantly, the ability to produce IFNγ and TNFα is not recovered by co-culture with iron replete dendritic cells, suggesting a T cell intrinsic alteration in functional memory formation. Altogether, these results isolate a critical effect of nutritional iron deficiency on T cell memory development and function.

During ontogeny, γδ T cells emerge from the thymus and directly seed peripheral tissues for in situ immunity. However, their functional role in humans has largely been defined from blood. Here, we analyzed the phenotype, transcriptome, function, and repertoire of human γδ T cells in blood and mucosal and lymphoid tissues from 176 donors across the life span, revealing distinct profiles in children compared with adults. In early life, clonally diverse Vδ1 subsets predominate across blood and tissues, comprising naïve and differentiated effector and tissue repair functions, whereas cytolytic Vδ2 subsets populate blood, spleen, and lungs. With age, Vδ1 and Vδ2 subsets exhibit clonal expansions and elevated cytolytic signatures, which are disseminated across sites. In adults, Vδ2 cells predominate in blood, whereas Vδ1 cells are enriched across tissues and express residency profiles. Thus, antigenic exposures over childhood drive the functional evolution and tissue compartmentalization of γδ T cells, leading to age-dependent roles in immunity.

Daptomycin is a last-line antibiotic commonly used to treat vancomycin-resistant Enterococci, but resistance evolves rapidly and further restricts already limited treatment options. While genetic determinants associated with clinical daptomycin resistance (DAP R ) have been described, information on factors affecting the speed of DAP R acquisition is limited. The multiple peptide resistance factor (MprF), a phosphatidylglycerol-modifying enzyme involved in cationic antimicrobial resistance, is linked to DAP R in pathogens such as methicillin-resistant Staphylococcus aureus. Since Enterococcus faecalis encodes two paralogs of mprF and clinical DAP R mutations do not map to mprF, we hypothesized that functional redundancy between the paralogs prevents mprF-mediated resistance and masks other evolutionary pathways to DAP R . Here, we performed in vitro evolution to DAP R in mprF mutant background. We discovered that the absence of mprF results in slowed DAP R evolution and is associated with inactivating mutations in ftsH, resulting in the depletion of the chaperone repressor HrcA. We also report that ftsH is essential in the parental, but not in the ΔmprF, strain where FtsH depletion results in growth impairment in the parental strain, a phenotype associated with reduced extracellular acidification and reduced ability for metabolic reduction. This presents FtsH and HrcA as enticing targets for developing anti-resistance strategies., (© 2024 The Authors. Molecular Microbiology published by John Wiley & Sons Ltd.)

Infections caused by nontuberculous mycobacteria have increased more than 50% in the past two decades and more than doubled in the elderly population. Mycobacterium abscessus (Mab), one of the most prevalent of these rapidly growing species, is intrinsically resistant to numerous antibiotics. Current standard-of-care treatments are not satisfactory, with high failure rate and notable adverse effects. We report here a potent anti-Mab compound from the flexible molecular framework afforded by conjugated oligoelectrolytes (COEs). A screen of structurally diverse, noncytotoxic COEs identified a lead compound, COE-PNH 2 , which was bactericidal against replicating, nonreplicating persisters and intracellular Mab.COE-PNH 2 had low propensity for resistance development, with a frequency of resistance below 1.25 × 10 -9 and showed no detectable resistance upon serial passaging. Mechanism of action studies were in line with COE-PNH 2 affecting the physical and functional integrity of the bacterial envelope and disrupting the mycomembrane and associated essential bioenergetic pathways. Moreover, COE-PNH 2 was well-tolerated and efficacious in a mouse model of Mab lung infection. This study highlights desirable in vitro and in vivo potency and safety index of this COE structure, which represents a promising anti-mycobacterial to tackle an unmet medical need.

The F 1 F O -ATP synthase engine is essential for viability and growth of nontuberculous mycobacteria (NTM) by providing the biological energy ATP and keeping ATP homeostasis under hypoxic stress conditions. Here, we report the discovery of the diarylquinoline TBAJ-5307 as a broad spectrum anti-NTM inhibitor, targeting the F O domain of the engine and preventing rotation and proton translocation. TBAJ-5307 is active at low nanomolar concentrations against fast- and slow-growing NTM as well as clinical isolates by depleting intrabacterial ATP. As demonstrated for the fast grower Mycobacterium abscessus, the compound is potent in vitro and in vivo, without inducing toxicity. Combining TBAJ-5307 with anti-NTM antibiotics or the oral tebipenem-avibactam pair showed attractive potentiation. Furthermore, the TBAJ-5307-tebipenem-avibactam cocktail kills the pathogen, suggesting a novel oral combination for the treatment of NTM lung infections., Competing Interests: Conflict of interest G. G., R. W. B., P. R., P. S.-L., and L. K. are inventors on the patent 23306555.6, which is related to the inhibitor described in this article. All other authors declare that they have no conflicts of interest with the contents of this article., (Copyright © 2024 The Authors. Published by Elsevier Inc. All rights reserved.)

Importance: New drugs are needed to combat multidrug-resistant tuberculosis. The electron transport chain (ETC) maintains the electrochemical potential across the cytoplasmic membrane and allows the production of ATP, the energy currency of any living cell. The mycobacterial engine F-ATP synthase catalyzes the formation of ATP and has come into focus as an attractive and rich drug target. Recent deep insights into these mycobacterial F 1 F O -ATP synthase elements opened the door for a renaissance of structure-based target identification and inhibitor design. In this study, we present the GaMF1.39 antimycobacterial compound, targeting the rotary subunit γ of the biological engine. The compound is bactericidal, inhibits infection ex vivo , and displays enhanced anti-tuberculosis activity in combination with ETC inhibitors, which promises new strategies to shorten tuberculosis chemotherapy., Competing Interests: G.G. and P.S.N. are inventors on the patent 10201911205R, which is related to the inhibitor described in this article.

Infancy and childhood are critical life stages for generating immune memory to protect against pathogens; however, the timing, location, and pathways for memory development in humans remain elusive. Here, we investigated T cells in mucosal sites, lymphoid tissues, and blood from 96 pediatric donors aged 0-10 years using phenotypic, functional, and transcriptomic profiling. Our results revealed that memory T cells preferentially localized in the intestines and lungs during infancy and accumulated more rapidly in mucosal sites compared with blood and lymphoid organs, consistent with site-specific antigen exposure. Early life mucosal memory T cells exhibit distinct functional capacities and stem-like transcriptional profiles. In later childhood, they progressively adopt proinflammatory functions and tissue-resident signatures, coincident with increased T cell receptor (TCR) clonal expansion in mucosal and lymphoid sites. Together, our findings identify staged development of memory T cells targeted to tissues during the formative years, informing how we might promote and monitor immunity in children., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2023 Elsevier Inc. All rights reserved.)

SARS-CoV-2 infection for most children results in mild or minimal symptoms, though in rare cases severe disease can develop, including a multisystem inflammatory syndrome (MIS-C) with myocarditis. Here, we present longitudinal profiling of immune responses during acute disease and following recovery in children who developed MIS-C, relative to children who experienced more typical symptoms of COVID-19. T cells in acute MIS-C exhibited transient signatures of activation, inflammation, and tissue residency which correlated with cardiac disease severity, while T cells in acute COVID-19 upregulated markers of follicular helper T cells for promoting antibody production. The resultant memory immune response in recovery showed increased frequencies of virus-specific memory T cells with pro-inflammatory functions in children with prior MIS-C compared to COVID-19 while both cohorts generated comparable antibody responses. Together our results reveal distinct effector and memory T cell responses in pediatric SARS-CoV-2 infection delineated by clinical syndrome, and a potential role for tissue-derived T cells in the immune pathology of systemic disease., (© 2023 Rybkina et al.)

The approval of the first-in-class antibacterial bedaquiline for tuberculosis marks a breakthrough in antituberculosis drug development. The drug inhibits mycobacterial respiration and represents the validation of a wholly different metabolic process as a druggable target space. In this review, we discuss the advances in the development of mycobacterial respiratory inhibitors, as well as the potential of applying this strategy to other pathogens. The non-fermentative nature of mycobacteria explains their vulnerability to respiration inhibition, and we caution that this strategy may not be equally effective in other organisms. Conversely, we also showcase fundamental studies that reveal ancillary functions of the respiratory pathway, which are crucial to some pathogens' virulence, drug susceptibility and fitness, introducing another perspective of targeting bacterial respiration as an antibiotic strategy., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 The Author(s). Published by Elsevier Ltd.. All rights reserved.)

Introduction: Tuberculosis (TB), caused by Mycobacterium tuberculosis (Mtb), is the most devastating bacterial disease. Multidrug-resistant Mtb strains are spreading worldwide, underscoring the need for new anti-TB targets and inhibitors. The respiratory chain complexes, including the cytochrome bd oxidase (cyt- bd ), have been identified as an attractive target for drug development. Recent novel structural and mechanistic insight as well as inhibitors of Mtb's cyt- bd brought this enzyme into the focus., Areas Covered: In this review, the authors describe conditions that stimulate the biogenesis of Mtb cyt- bd , its structural-, mechanistic-, and substrate-binding traits. They discuss the present Mtb cyt- bd inhibitors, novel targets within the enzyme and structure activity relationship features that are required for mycobacterial cyt- bd inhibition and augment their understanding on improving the potency of cyt- bd inhibitors., Expert Opinion: A deeper structure-mechanistic understanding of Mtb's cyt- bd is a prerequisite for in silico efforts to: (i) identify pathogen specific targets for the design of novel nontoxic hit molecules, forming the platform for the development of new leads, (ii) design mechanism of action studies, (iii) perform medicinal chemistry of existing inhibitors to improve their potency and pharmacokinetic/-dynamic properties. Phase studies with such optimized cyt- bd inhibitors in combination with anti-TB compounds targeting the oxidative phosphorylation pathway is recommended.

The mycobacterial cytochrome bcc:aa 3 complex deserves the name "supercomplex" since it combines three cytochrome oxidases-cytochrome bc , cytochrome c , and cytochrome aa 3 -into one supramolecular machine and performs electron transfer for the reduction of oxygen to water and proton transport to generate the proton motive force for ATP synthesis. Thus, the bcc:aa 3 complex represents a valid drug target for Mycobacterium tuberculosis infections. The production and purification of an entire M. tuberculosis cytochrome bcc:aa 3 are fundamental for biochemical and structural characterization of this supercomplex, paving the way for new inhibitor targets and molecules. Here, we produced and purified the entire and active M. tuberculosis cyt- bcc:aa 3 oxidase, as demonstrated by the different heme spectra and an oxygen consumption assay. The resolved M. tuberculosis cyt- bcc:aa 3 cryo-electron microscopy structure reveals a dimer with its functional domains involved in electron, proton, oxygen transfer, and oxygen reduction. The structure shows the two cytochrome c I c II head domains of the dimer, the counterpart of the soluble mitochondrial cytochrome c , in a so-called "closed state," in which electrons are translocated from the bcc to the aa 3 domain. The structural and mechanistic insights provided the basis for a virtual screening campaign that identified a potent M. tuberculosis cyt- bcc:aa 3 inhibitor, cyt Mycc 1. cyt Mycc 1 targets the mycobacterium-specific α3-helix of cytochrome c I and interferes with oxygen consumption by interrupting electron translocation via the c I c II head. The successful identification of a new cyt- bcc:aa 3 inhibitor demonstrates the potential of a structure-mechanism-based approach for novel compound development., Competing Interests: The authors declare no conflict of interest.

The bioenergetic mechanisms by which Mycobacterium tuberculosis survives hypoxia are poorly understood. Current models assume that the bacterium shifts to an alternate electron acceptor or fermentation to maintain membrane potential and ATP synthesis. Counterintuitively, we find here that oxygen itself is the principal terminal electron acceptor during hypoxic dormancy. M. tuberculosis can metabolize oxygen efficiently at least two orders of magnitude below the concentration predicted to occur in hypoxic lung granulomas. Despite a difference in apparent affinity for oxygen, both the cytochrome bcc:aa 3 and cytochrome bd oxidase respiratory branches are required for hypoxic respiration. Simultaneous inhibition of both oxidases blocks oxygen consumption, reduces ATP levels, and kills M. tuberculosis under hypoxia. The capacity of mycobacteria to scavenge trace levels of oxygen, coupled with the absence of complex regulatory mechanisms to achieve hierarchal control of the terminal oxidases, may be a key determinant of long-term M. tuberculosis survival in hypoxic lung granulomas., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2023 The Author(s). Published by Elsevier Inc. All rights reserved.)

Background: Mycobacterium abscessus is a non-tuberculous mycobacterium (NTM) that causes chronic pulmonary infections. Because of its extensive innate resistance to numerous antibiotics, treatment options are limited, often resulting in poor clinical outcomes. Current treatment regimens usually involve a combination of antibiotics, with clarithromycin being the cornerstone of NTM treatments., Objectives: To identify drug candidates that exhibit synergistic activity with clarithromycin against M. abscessus ., Methods: We performed cell-based phenotypic screening of a compound library against M. abscessus induced to become resistant to clarithromycin. Furthermore, we evaluated the toxicity and efficacy of the top compound in a zebrafish embryo infection model., Results: The screen revealed rifaximin as a clarithromycin potentiator. The combination of rifaximin and clarithromycin was synergistic and bactericidal in vitro and potent in the zebrafish model., Conclusions: The data indicate that the rifaximin/clarithromycin combination is promising to effectively treat pulmonary NTM infections., (© The Author(s) 2023. Published by Oxford University Press on behalf of British Society for Antimicrobial Chemotherapy.)

New antimicrobials are urgently needed to combat Gram-negative bacteria, particularly multi-drug resistant (MDR) and phenotypically resistant biofilm species. At present, only sequence-defined alpha-peptides (e.g. polymyxin B) can selectively target Gram-negative bacterial lipopolysaccharides. We show that a copolymer, without a defined sequence, shows good potency against MDR Gram-negative bacteria including its biofilm form. The tapered blocky co-beta-peptide with controlled N-terminal hydrophobicity (#4) has strong interaction with the Gram-negative bacterial lipopolysaccharides via its backbone through electrostatic and hydrogen bonding interactions but not the Gram-positive bacterial and mammalian cell membranes so that this copolymer is non-toxic to these two latter cell types. The new #4 co-beta-peptide selectively kills Gram-negative bacteria with low cytotoxicity both in vitro and in a mouse biofilm wound infection model. This strategy provides a new concept for the design of Gram-negative selective antimicrobial peptidomimetics against MDR and biofilm species., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier Ltd. All rights reserved.)

The antimicrobial resistance crisis is a global health issue requiring discovery and development of novel therapeutics. However, conventional screening of natural products or synthetic chemical libraries is uncertain. Combination therapy using approved antibiotics with inhibitors targeting innate resistance mechanisms provides an alternative strategy to develop potent therapeutics. This review discusses the chemical structures of effective β-lactamase inhibitors, outer membrane permeabilizers, and efflux pump inhibitors that act as adjuvant molecules of classical antibiotics. Rational design of the chemical structures of adjuvants will provide methods to impart or restore efficacy to classical antibiotics for inherently antibiotic-resistant bacteria. As many bacteria have multiple resistance pathways, adjuvant molecules simultaneously targeting multiple pathways are promising approaches to combat multidrug-resistant bacterial infections., Competing Interests: The authors declare no competing financial interest., (© 2023 The Authors. Published by American Chemical Society.)

New therapeutic concepts are critically needed for carbapenem-resistant Pseudomonas aeruginosa, an opportunistic pathogen particularly recalcitrant to antibiotics. The screening of around 230,000 small molecules yielded a very low hit rate of 0.002% after triaging for known antibiotics. The only novel hit that stood out was the antimetabolite oxythiamine. Oxythiamine is a known transketolase inhibitor in eukaryotic cells, but its antibacterial potency has not been reported. Metabolic and transcriptomic analyses indicated that oxythiamine is intracellularly converted to oxythiamine pyrophosphate and subsequently inhibits several vitamin-B1-dependent enzymes, sensitizing the bacteria to several antibiotic and non-antibiotic drugs such as tetracyclines, 5-fluorouracil, and auranofin. The positive interaction between 5-fluorouracil and oxythiamine was confirmed in a murine ocular infection model, indicating relevance during infection. Together, this study revealed a system-level significance of thiamine metabolism perturbation that sensitizes P. aeruginosa to multiple small molecules, a property that could inform on the development of a rational drug combination., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2022 Elsevier Ltd. All rights reserved.)

Introduction: Currently, a pediatric mental and behavioral health crisis exists, driven by increasing stressors among children coupled with a paucity of psychiatric providers who treat children. Pediatric primary care providers can play a critical role in filling this gap, yet trainees feel uncomfortable screening for, identifying, and managing mental and behavioral health conditions among their patients. Thus, expanding training for pediatricians in this domain is critical., Methods: We created a longitudinal integrated mental and behavioral health curriculum for pediatric residents at NewYork-Presbyterian/Columbia University Irving Medical Center with a logic model contextualizing outpatient pediatric care as a framework for the development and planned evaluation. We devised a comprehensive set of materials, with presentations on topics including attention deficit hyperactivity disorder and anxiety disorders. Workflows and escalation pathways promoting collaboration among interdisciplinary providers were implemented. We evaluated residents' and faculty members' participation in the curriculum and their perception of curricular gaps., Results: Approximately 155 pediatric residents participated in the curriculum from 2017 to 2021, reflecting robust curricular exposure. Few residents and no preceptors perceived mental and behavioral health as a curricular gap., Discussion: Our curriculum is feasible and can be adapted to a variety of educational settings. Its use of a logic model for development, implementation, and ongoing evaluation grounds the curriculum in educational theory and can address curricular gaps. The framework can be adapted to suit the needs of other institutions' educational and practice settings and equip pediatric trainees with the skills to promote patient mental health and well-being., (© 2022 Meyers et al.)

The treatment of leprosy is long and complex, benefiting from the development of sterilizing, rapidly-acting drugs. Reductive evolution made Mycobacterium leprae exquisitely sensitive to Telacebec, a phase 2 drug candidate for tuberculosis . The unprecedented potency of Telacebec against M. leprae warrants further validation in clinical trials.

Supracondylar humerus fractures (ScHF) account for 60% of fractures of the elbow region in children. We assessed the relationship between neurovascular complications and the degree of fracture displacement as rated on the basis of modified Gartland classification. Moreover, we aimed to evaluate predisposing factors, e.g., age and gender, and outcomes of neurovascular complications in ScHF. Between 2004 and 2019, we treated 329 patients with ScHF at the Department of Traumatology and Orthopedics of the Upper Silesian Child Centre, Katowice, Poland. Mean age of patients (189 boys and 140 girls) was 7.2 years (Confidence interval: 6.89, 7.45). Undisplaced fractures were treated conservatively with a cast. Displaced fractures were managed by closed reduction and percutaneous Kirschner wire fixation using two pins inserted laterally. We retrospectively assessed the number of neurovascular lesions at baseline and recorded any iatrogenic injury resulting from the surgical intervention. Acute neurovascular lesions occurred in 44 of 329 ScHF patients (13.4%). The incidence of accompanying neurovascular injuries was positively associated with the severity of fracture displacement characterized by Gartland score. Vascular injuries occurred mainly in Gartland type IV ScHF, while nerve lesions occurred in both Gartland type III and IV ScHF. We noted a significantly higher mean Gartland score and mean age at injury in the group of children suffering from neurovascular injuries when compared to those in the group without such injuries ( p = 0.045 and p = 0.04, respectively). We observed no secondary nerve lesions after surgical treatment. For the treatment of ScHF in children, we recommend closed reduction and stabilization of displaced fractures with K-wires inserted percutaneously from the lateral aspect of the upper arm. We advocate vessel exploration in case of absent distal pulses after closed reduction but do not consider primary nerve exploration necessary, unless a complete primary sensomotoric nerve lesion is present.

Introduction: Tuberculosis is an infectious disease that affected more than 50 million people and killed 6.7 million patients in the past 5 years alone. Additionally, rising incidence of treatment resistance threatens the global effort to eradicate this disease. With limited options available, additional novel antibiotics are needed for the treatment of multidrug-resistant tuberculosis (MDR-TB). Telacebec is a first-in-class antibiotic that targets the pathogen's energy metabolism., Areas Covered: This paper provides an overview of the recent progress in the development and testing of telacebec. We discuss published clinical data and examine the design and setup of its clinical trials. We also offer insights on the therapeutic potential of telacebec and aspects of which should be evaluated in the future., Expert Opinion: The first phase 2a trial showed a correlation between dosage and bacterial load in patient sputum, which should be confirmed using a direct measurement method such as colony-forming unit counting. Its clinical efficacy, favorable pharmacokinetic properties, low arrhythmogenic risk, and activity against MDR-TB strains make telacebec a suitable candidate for further development. Future clinical testing in combination with approved second-line drugs will reveal its full potential against MDR-TB. Considering recent preclinical studies, we also recommend initiating clinical trials for Buruli ulcer and leprosy.

The oxidative phosphorylation (OxPhos) pathway has emerged as an attractive pathway for the development of anti-mycobacterial drugs. The OxPhos pathway is essential for ATP resynthesis and maintenance of the electrochemical transmembrane gradient. The bioenergetic parameters of the pathway such as oxygen consumption rate and ATP levels are quantifiable using current technology. Measuring these parameters are useful tools to gauge rapidly the impact of drug candidates on their capacity to inhibit the OxPhos pathway in Mycobacterium ulcerans., (© 2022. The Author(s), under exclusive license to Springer Science+Business Media, LLC, part of Springer Nature.)

Generation and characterization of drug resistant mutants is a powerful tool in antimicrobial drug discovery for identification of the molecular target of an investigational drug candidate. The method is relatively simple to be conducted in a classical microbiology laboratory. Its value has been augmented by the employment of next generation sequencing techniques to characterize single-nucleotide polymorphisms associated with drug resistance. Determination of the frequency of emergence of resistance to drug candidates also provides insights into their usefulness for clinical application. In addition to the generation of drug resistant mutants, we describe a direct method to determine the minimum inhibitory concentration of a drug candidate against Mycobacterium ulcerans., (© 2022. The Author(s), under exclusive license to Springer Science+Business Media, LLC, part of Springer Nature.)

The growing prevalence of antimicrobial drug resistance in pathogenic bacteria is a critical threat to global health. Conventional antibiotics still play a crucial role in treating bacterial infections, but the emergence and spread of antibiotic-resistant micro-organisms are rapidly eroding their usefulness. Cationic polymers, which target bacterial membranes, are thought to be the last frontier in antibacterial development. This class of molecules possesses several advantages including a low propensity for emergence of resistance and rapid bactericidal effect. This review surveys the structure-activity of advanced antimicrobial cationic polymers, including poly(α-amino acids), β-peptides, polycarbonates, star polymers and main-chain cationic polymers, with low toxicity and high selectivity to potentially become useful for real applications. Their uses as potentiating adjuvants to overcome bacterial membrane-related resistance mechanisms and as antibiofilm agents are also covered. The review is intended to provide valuable information for design and development of cationic polymers as antimicrobial and antibiofilm agents for translational applications., Competing Interests: The authors declare no competing financial interests., (This journal is © The Royal Society of Chemistry.)

The antituberculosis drug telacebec is ineffective against Mycobacterium abscessus. A recent study suggested that TB47, a telacebec analogue, potentiated the efficacy of clofazimine against M. abscessus. Here, we report that TB47 not only is ineffective against M. abscessus in vitro but also does not potentiate the activity of clofazimine.

Background: Acetabular fractures are rare in children, constituting approximately 1-4.6% of all paediatric fractures., Material and Methods: Nine patients (4 girls and 5 boys) with a mean age of 14.5 years (range, 12-16.5 years) were treated due to acetabular fractures between 2000 and 2020. Most of the fractures were caused by road accidents. Before the patients were qualified for treatment, they underwent a CT scan (all patients) and an MRI scan (8 patients) of the hip. Surgical treatment was used in 6 patients while the other 2 were managed conservatively., Results: Mean follow-up period was 6.2 years (range, 2-10 years). Bone union was achieved in all patients after 76 days on average (range, 65-90 days). The mean Merle d'Aubigne score at 12 months after treatment was 17.1 points (range, 13-18 points)., Conclusions: The treatment of acetabular fractures in paediatric patients during or after puberty may be the same as in adults. An MRI scan of the hip joint is recommended for assessing the fracture, particularly the triradiate cartilage.

The development of cytochrome bd oxidase (cyt- bd ) inhibitors are needed for comprehensive termination of energy production in Mycobacterium tuberculosis (Mtb) to treat tuberculosis infections. Herein, we report on the structure-activity-relationships (SAR) of 22 new N -phenethyl-quinazolin-4-yl-amines that target cyt- bd . Our focused set of compounds was synthesized and screened against three mycobacterial strains: Mycobacterium bovis BCG, Mycobacterium tuberculosis H37Rv and the clinical isolate Mycobacterium tuberculosis N0145 with and without the cytochrome bcc:aa 3 inhibitor Q203 in an ATP depletion assay. Two compounds, 12a and 19a , were more active against all three strains than the naturally derived cyt- bd inhibitor aurachin D., Competing Interests: Conflicts of Interest: Hsiri Therapeutics has licensed this technology. M.J.M. and G.C.M. own equity in Hsiri. M.J.M. is CSO of Hsiri. G.C.M. and K.P. are consultants to Hsiri. Hsiri Therapeutics did not fund this study and was not involved in study design or interpretation.

Background: Occipital condyle fractures (OCFs) in patients before 18 years of age are rare. Classifications of OCF are based on the CT images of the cranio-cervical junction (CCJ) and MRI. The Anderson-Montesano and Tuli classifications are the types which are most commonly used in these cases. Classification of OCFs allows the implementation of OCF treatment. The aim of this study was to evaluate the effectiveness of using the OCF classification in pediatric patients based on the analysis of our own cases., Methods: During the years 2013-2020, 6 pediatric patients with OCFs, aged 14-18, have been treated. Two patients with unstable fracture III according to Anderson-Montesano and IIB according to Tuli were treated with the halo-vest. Additionally, one patient presenting neurological symptoms and with an associated C1 fracture was qualified for the halo-vest stabilization as well. The other patients were treated with a Minerva collar. We evaluated the results 6 months after completing the OCF treatment using the Neck Disability Index (NDI) and SF-36 questionnaires. Confidence intervals for the mean values were verified using the MeanCI function (from the R library DescTools) for both classical and bootstrap methods., Results: Based on NDI results, we have obtained in our patients an average of 4.33/45 points (2-11) and 9.62% (4.4-24.4). Based on the SF-36 questionnaire, we obtained an average of 88.62% (47.41-99.44)., Conclusion: The Anderson-Montesano and Tuli's classifications of OCF can be used to assess the stability of OCF in adolescents, but both classifications should be used simultaneously. CT and MR imaging should be used in diagnosing OCFs, whereas CT allows assessing therapeutic outcomes in OCF., (© 2021. The Author(s).)

Zwitterionic polymers are classical antifouling polymers but they require specialized monomers that have cationic and anionic charges integrated into a single monomer. Herein, we show that pseudo-zwitterionic copolymers synthesized from a mixture of 2 monomers each having a single opposite polarity has excellent antibiofilm efficacy. We have discovered a new mixed-charge copolymer brush (#1-A) synthesized from 2 oppositely charged monomers, the anionic SPM (3-Sulfopropyl methacrylate) and the cationic AMPTMA ((3-Acrylamidopropyl) trimethylammonium chloride), that achieves broad spectrum in vitro antibiofilm effect of greater than 99% reductions against all six Gram-positive and Gram-negative bacteria tested. In the murine subcutaneous wound catheter infection models, the #1-A has good long-term anti-biofilm efficacy against MRSA and Pseudomonas aeruginosa of 3.41 and 3.19 orders respectively, outperforming previous mixed-charge copolymer coatings. We discovered a new method to choose the cationic/anionic pair combination to form the best antibiofilm copolymer brush coating by exploiting the solution polymerization kinetics disparity between the cationic and anionic monomers. We also showed that #1-A is softer and has higher hydration than the classical zwitterionic polymer. This study shows the possibility of achieving potent antibiofilm efficacy by combining readily available opposite singly charged monomers., (Copyright © 2021 Elsevier Ltd. All rights reserved.)

Antimicrobial resistance is a global threat. In addition to the emergence of resistance to last resort drugs, bacteria escape antibiotics killing by forming complex biofilms. Strategies to tackle antibiotic resistance as well as biofilms are urgently needed. Wall teichoic acid (WTA), a generic anionic glycopolymer present on the cell surface of many Gram-positive bacteria, has been proposed as a possible therapeutic target, but its druggability remains to be demonstrated. Here we report a cationic glycosylated block co-β-peptide that binds to WTA. By doing so, the co-β-peptide not only inhibits biofilm formation, it also disperses preformed biofilms in several Gram-positive bacteria and resensitizes methicillin-resistant Staphylococcus aureus to oxacillin. The cationic block of the co-β-peptide physically interacts with the anionic WTA within the cell envelope, whereas the glycosylated block forms a nonfouling corona around the bacteria. This reduces physical interaction between bacteria-substrate and bacteria-biofilm matrix, leading to biofilm inhibition and dispersal. The WTA-targeting co-β-peptide is a promising lead for the future development of broad-spectrum anti-biofilm strategies against Gram-positive bacteria.

In the setting of COVID-19, pediatric primary care in New York City faced multiple challenges, requiring large-scale practice reorganization. We used quality improvement principles to implement changes to care delivery rapidly., Methods: Plan-do-study-act cycles were used, based on primary drivers of consolidation, reorganization of in-person and urgent care, telehealth expansion, patient outreach, mental health linkages, team communication, and safety., Results: The average visit volume in pediatrics decreased from 662 per week to 370. Telehealth visits increased from 2 to 140 per week, whereas urgent in-person visits decreased from 350 to 8 per week. Adolescent visits decreased from 57 to 46 per week. Newborn Clinic visits increased from 37 per week to 54. Show rates increased significantly for pediatrics and adolescent ( P = 0.003 and P = 0.038, respectively)., Conclusions: Quality improvement methodology allowed for the consolidation of pediatric primary care practices during the first wave of the COVID-19 pandemic, ensuring care for patients while prioritizing safety, evidence-based practices, and available resources., Competing Interests: The authors have no financial interest to declare in relation to the content of this article., (Copyright © 2020 the Author(s). Published by Wolters Kluwer Health, Inc.)

Objective: Dermatologic complaints are common in outpatient pediatrics. However, pediatric dermatology specialty care can be difficult to access. We aimed to test the feasibility of co-locating dermatology services within primary care and increase the proportion of patients treated for basic skin complaints within the medical home while decreasing wait times., Methods: The Rapid Assessment of Skin Health (RASH) clinic was created within a hospital-based primary care clinic in 11/2013. The clinic was staffed by 2 pediatricians trained in the dermatology department and supported with specialist advice as needed. Referral volume and wait times to dermatology and RASH clinic were tracked for visits between 11/1/12 and 10/31/18. A chart review was also conducted on a subset of RASH clinic visits. Primary care providers (PCPs) were surveyed about their experiences., Results: Fifty-eight percent of patients referred for a dermatologic complaint were scheduled in RASH clinic. Wait times for new patient appointments in RASH clinic were significantly shorter than for new dermatology appointments in the previous 12 months (mean 36 days vs 65 days, P < .001). The monthly number of referrals to dermatology also decreased significantly after the RASH clinic opened (24/month vs 12/month, P < .001). Ten percent of RASH patients were referred on to dermatology. In a survey of PCPs (N = 67), 76% said the RASH clinic was "extremely/very helpful.", Conclusions: Providing dermatologic care to low or moderate complexity patients within the medical home is feasible and leads to better access to care. This innovative model could be spread to other clinics and subspecialties., (Copyright © 2020 Academic Pediatric Association. Published by Elsevier Inc. All rights reserved.)

GTPase high f requency of l ysogenization X (HflX) is highly conserved in prokaryotes and acts as a ribosome-splitting factor as part of the heat shock response in Escherichia coli. Here we report that HflX produced by slow-growing Mycobacterium bovis bacillus Calmette-Guérin (BCG) is a GTPase that plays a critical role in the pathogen's transition to a nonreplicating, drug-tolerant state in response to hypoxia. Indeed, HflX-deficient M. bovis BCG (KO) replicated markedly faster in the microaerophilic phase of a hypoxia model that resulted in premature entry into dormancy. The KO mutant displayed hallmarks of nonreplicating mycobacteria, including phenotypic drug resistance, altered morphology, low intracellular ATP levels, and overexpression of Dormancy (Dos) regulon proteins. Mice nasally infected with HflX KO mutant displayed increased bacterial burden in the lungs, spleen, and lymph nodes during the chronic phase of infection, consistent with the higher replication rate observed in vitro in microaerophilic conditions. Unlike fast growing mycobacteria, M. bovis BCG HlfX was not involved in antibiotic resistance under aerobic growth. Proteomics, pull-down, and ribo-sequencing approaches supported that mycobacterial HflX is a ribosome-binding protein that controls translational activity of the cell. With HflX fully conserved between M. bovis BCG and M. tuberculosis , our work provides further insights into the molecular mechanisms deployed by pathogenic mycobacteria to adapt to their hypoxic microenvironment., Competing Interests: The authors declare no competing interest.

Antimicrobial peptides that target the integrity of bacterial envelopes can eradicate pathogens with little development of resistance, but they often inflict nonselective toxicity toward mammalian cells. The prevailing approach to optimize the selectivity of cationic peptides has been to modify their composition. Instead, we invent a new generation of broad-spectrum antibacterial nanoconstructs with negligible mammalian cell toxicity through a competitive displacement of counter polyanions from the complementary polycations. The nanoconstruct, which has a highly cationic Au nanoparticles (NPs) core shielded by polymeric counterions, is inert in nonbacterial environments. When exposed to negatively charged bacterial envelopes, this construct sheds its polyanions, triggering a cationic Au NP/bacterial membrane interaction that rapidly kills Gram-positive and Gram-negative bacteria. The anionic charge and hydrophilicity of the polyanion provides charge neutralization for the peptide-decorated Au NP core, but it is also bacteria-displaceable. These results provide a foundation for the development of other cationic particles and polymeric counterion combinations with potent antimicrobial activity without toxicity.

Cationic polymers are promising antibacterial agents because bacteria have a low propensity to develop resistance against them, but they usually have low biocompatibility because of their hydrophobic moieties. Herein, we report a new biodegradable and biocompatible chitosan-derived cationic antibacterial polymer, 2,6-diamino chitosan (2,6-DAC). 2,6-DAC shows excellent broad-spectrum antimicrobial activity with minimum inhibitory concentrations (MICs) of 8-32 μg/mL against clinically relevant and multidrug-resistant (MDR) bacteria including Listeria monocytogenes , Staphylococcus aureus , Escherichia coli , Klebsiella pneumoniae , Pseudomonas aeruginosa , and Acinetobacter baumannii . Furthermore, 2,6-DAC shows an excellent synergistic effect with various clinically relevant antibiotics proved by decreasing the MICs of the antibiotics against MDR A. baumannii and methicillin-resistant Staphylococcus aureus to <1 μg/mL. In vivo biocompatibility of 2,6-DAC is proved by a dosage of 100 mg/kg compound via oral administration and 25 mg/kg compound via intraperitoneal injection to mice; 2,6-DAC does not cause any weight loss and any significant change in liver and kidney biomarkers or the important blood electrolytes. The combinations of 2,6-DAC together with novobiocin and rifampicin show >2.4 log 10 reduction of A. baumannii in murine intraperitoneal and lung infection models. The novel chitosan derivative, 2,6-DAC, can be utilized as a biocompatible broad-spectrum cationic antimicrobial agent alone or in synergistic combination with various antibiotics.